Many cellular processes involve transformations and sustenance of mechanical forces. Forces arise as input, output and signals in cellular machines, e.g., in ATP synthase, in motor proteins, and in integrin cell surface receptors. Proteins need to sustain the considerable forces arising in cells, e.g., in the case of muscle proteins and the extracellular matrix. For several years our group has focused its research on the mechanical stability of proteins under the action of naturally arising forces, employing its well established methodology of steered molecular dynamics simulations. We plan to continue collaborations with several experimental laboratories to take advantage of research opportunities that have arisen through recent discoveries. We propose to study the architecture of the giant muscle protein titin and related proteins. Within muscle fibers the ends of two titin molecules are glued to each other through a ligand protein called telethonin by means of beta-strand cross-linking, which provides a novel mechanism for stabilizing a ligand-receptor complex. We seek to elucidate the structure-function relationship of this complex. In a close collaboration with a crystallographic laboratory new structures of hitherto inaccessible titin domains will be solved and investigated computationally. Investigation of Myomesin, a miniature of titin and cross-linking muscle thick filaments, will provide understanding on the structural design essential for a minimized elastic protein. The ability of ankyrin, found as linker proteins in many mammalian tissues, to act as a non-entropic spring will be characterized with a particular application to inner ear mechanics. The effects of mutations related to hereditary deafness on the stability and elasticity of cadherin will be studied as well. Finally the mechanical properties of two classic gene regulators, lac and lambda repressors, will be characterized by investigating the interaction of DNA loops with the proteins through multiscale modeling. The proposed effort is based on a systematic development of tools for large-scale molecular modeling as well as on published and preliminary studies that have already progressed considerably towards the ambitious goals stated in the proposal. [unreadable] [unreadable] [unreadable]